Kits and methods for assessing skin health

ABSTRACT

The invention relates to kits and methods for assessing skin health for a human and the human&#39;s susceptibility to skin disorders. The methods involve assessing occurrence in the human&#39;s genome of one or more polymorphisms (e.g., single nucleotide polymorphisms) that occur in one or more genes associated disclosed herein and that are associated with a disorder in humans. Preferred assessment and scoring methods are disclosed, as are kits for performing the methods.

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application is a continuation-in-part of InternationalApplication No. PCT/US02/10682 (filed 5 Apr. 2002) and of U.S. patentapplication Ser. No. 09/826,522 (filed 5 Apr. 2001), and is entitled topriority pursuant to 35 U.S.C. §119(e) to U.S. provisional patentapplications No. 60/289,169 (filed 7 May 2001), No. 60/350,517 (filed 22Oct. 2001), No. 60/335,426 (filed 24 Oct. 2001), and No. 60/336/815(filed 5 Dec. 2001).

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT

[0002] Not applicable.

REFERENCE TO A MICROFICHE APPENDIX

[0003] Not applicable.

BACKGROUND OF THE INVENTION

[0004] Skin is the largest and most visible organ of the human body, andis also among the tissues most exposed to environmental stresses,hazards, and pathogens. Skin is a multi-layered tissue, primarilycomposed of the epidermis and dermis, and includes several accessorystructures, such as sweat glands, sebaceous glands, and hair follicles.Skin serves many functions. For example, it is a protective barrier toexternal insults (e.g., heat, chemicals, bacteria), is involved inthermoregulation, inhibits dehydration, and performs sensory functions.Skin is also a bioreactor that produces various hormones and lipids thatenter the body's circulation. A variety of immune cells function in skinas a first line of defense against bacterial or viral invasion and tomaintain immune surveillance in skin and nearby body tissues. For thesereasons, establishment and maintenance of good skin health is importantto human health.

[0005] Skin health is also important for aesthetic reasons. Many peopleare deeply concerned about the appearance of their skin. A healthy skinappearance is maintained by a combination of cleaning, nutrition, andapplication of therapeutic and cosmetic products. However, overuse ofskin care products can degrade skin health and appearance. Often,individuals employ trial-and-error techniques to identify skin careproducts (and doses thereof) that produce a desirable skin appearance.More precise methods are needed for identifying compositions (andsuitable amounts of such compositions) that will enhance the health andappearance of an individual's skin. These methods would preferably betailored to identify useful compositions and dosages for individuals.The present invention satisfies this need.

[0006] Many skin disorders can be alleviated, inhibited, or evenprevented by maintaining a high degree of skin health or by timelyintervention with appropriate skin-affecting agents. For example, suchintervention can include consuming or topically applying skin careproducts, modulating sun exposure, adjusting diet, consuming nutritionalor pharmaceutical products known to be effective against skin disorders,and undergoing heightened medical monitoring. These changes are oftennot made, owing to the expense or inconvenience of the changes and anindividual's subjective belief that he or she is not at high risk forskin disorders. Improved assessment of skin health can help to identifyindividuals at risk for developing skin disorders and permit moreinformed decisions to be made regarding whether lifestyle changes orother interventions are justified.

[0007] Many human genes occur in a variety of forms which differ in atleast minor ways. Heterogeneity in human genes is believed to havearisen, in part, from minor, non-fatal mutations that have occurred inthe genome over time. In some instances, differences between alternativeforms of a gene are manifested as differences in the amino acid sequenceof a protein encoded by the gene. Some amino acid sequence differencescan alter the reactivity or substrate specificity of the protein.Differences between alternative forms of a gene can also affect thedegree to which (if at all) the gene is expressed. However, manyheterogeneities that occur in human genes appear not to be correlatedwith any particular phenotype. Known heterogeneities include, forexample, single nucleotide polymorphisms (i.e., alternative forms of agene having a difference at a single nucleotide residue). Other knownpolymorphic forms include those in which the sequence of larger (e.g.,2-1000 residues) portions of a gene exhibits numerous sequencedifferences and those which differ by the presence or absence of portionof a gene.

[0008] Numerous disorders and physiological states have been correlatedwith occurrence of one or more alternative forms of an individual genein the genome of a human who exhibits the disorder or physiologicalstate. For example, Kimura et al. (2000, Am. J. Ophthalmol. 130:769-773)discloses an association between occurrence of a SNP of the manganesesuperoxide dismutase gene and a form of macular degeneration.

[0009] Associations between individual disorders and individual geneticpolymorphisms are known. However, disorders can usually result frompolymorphisms in any of a relatively large number of genes, and as aresult, assessing the polymorphic form(s) of any single gene that occurin a human's genome is usually not predictive of the likelihood that thehuman will develop the disorder.

[0010] A need remains for a method of assessing an individual's skinhealth or predisposition to develop skin disorders. Such assessmentcould be used to identify types and amounts of therapeutic, inhibitory,or preventive compositions or interventions that can be used toalleviate, inhibit, or prevent skin disorders. The invention satisfiesthese needs.

BRIEF SUMMARY OF THE INVENTION

[0011] The invention relates to a method of assessing skin health in ahuman. The method comprises assessing occurrence in the human's genomeof disorder-associated polymorphisms in at least two (three, four, five,six, eight, ten, fifteen, or even all) genes selected from the groupconsisting of

[0012] genes which encode an enzyme that catalyzes conversion of a toxicoxygen species to a less toxic oxygen species;

[0013] genes which encode a protein that provides protection againstoxidative stress;

[0014] genes which encode a protein that induces production of a toxicoxygen species;

[0015] genes which encode a protein that indirectly affects oxidativestress;

[0016] genes which encode a protein for which the level of expression ofthe protein is associated with oxidative stress;

[0017] genes which encode a component of the human DNA repair system;and

[0018] genes which encode a protein associated with production of atoxic oxygen species by a macrophage or polymorphonuclear neutrophilicgranulocyte.

[0019] It has been discovered that this method is particularly usefulfor assessing skin health when the genes are selected from the groupconsisting of

[0020] a) the gene which encodes mitochondrial manganese superoxidedismutase (MnSOD);

[0021] b) the gene which encodes cytoplasmic copper/zinc superoxidedismutase (CZSOD);

[0022] c) the gene which encodes catalase;

[0023] d) the gene which encodes human glutathione peroxidase (hGPX1);

[0024] e) the gene which encodes glutathione S transferase P1 (GSTP1)

[0025] f) the gene which encodes NAD(P)H:quinone oxidoreductase;

[0026] g) the gene which encodes epoxide hydrolase;

[0027] h) the gene which encodes tumor necrosis factor alpha(TNF-alpha);

[0028] i) the gene which encodes NADH/NADPH oxidase p22 subunit (thephox gene);

[0029] j) the gene which encodes nitric oxide synthase;

[0030] k) the gene which encodes cytochrome P450;

[0031] l) the gene which encodes matrix metalloproteinase 1 (MMP-1); and

[0032] m) the gene which encodes profilagrin.

[0033] Occurrence of a disorder-associated polymorphism in any of thesegenes is an indication that the human has poorer skin health than ahuman whose genome does not comprise the disorder-associatedpolymorphism, and occurrence of a plurality of disorder-associatedpolymorphisms is an indication that the human has even poorer skinhealth than a human whose genome comprises only one of thedisorder-associated polymorphisms (and greater still than an individualwhose genome does not comprise one of these disorder-associatedpolymorphisms).

[0034] Substantially the same method can be used to assess theadvisability that a human should employ a skin care product, such as onecomprising a skin protective ingredient or a vitamin (e.g., one ofvitamins C and E). When the method is used to assess the advisabilitythat a human should employ a skin care product, occurrence of one ormore disorder-associated polymorphisms in any of genes a)-l) is anindication that it is more advisable for the human to use the productthan when the individual's genome does not comprise disorder-associatedpolymorphisms in any of these genes.

[0035] For example, occurrence of at least two (three, four, five, six,eight, ten, or fifteen or more) disorder-associated polymorphisms can beassessed, where the polymorphisms are selected from the group consistingof

[0036] A) a polymorphism in the open reading frame encodingmitochondrial MnSOD;

[0037] B) a polymorphism in the open reading frame encoding cytoplasmicCZSOD;

[0038] C) a polymorphism in the promoter region of the gene encodingcatalase;

[0039] D) a polymorphism in the open reading frame of the hGPX1 gene;

[0040] E) a polymorphism in the open reading frame encoding glutathioneS transferase P1 (GSTP1);

[0041] F) a polymorphism in the open reading frame encodingNAD(P)H:quinone oxidoreductase;

[0042] G) a polymorphism in the open reading frame encoding epoxidehydrolase;

[0043] H) a polymorphism in the promoter region of the gene encodingTNF-alpha;

[0044] I) a polymorphism in the open reading frame of the phox gene;

[0045] J) a polymorphism in the open reading frame encoding nitric oxidesynthase;

[0046] K) a polymorphism in the 5′ flanking region of the gene encodingcytochrome P450; and

[0047] L) a polymorphism in the promoter region of the gene encodingMMP-1.

[0048] Preferably, occurrence of all known polymorphisms at individualsites (e.g., both of two known alternative forms or all three forms of apolymorphism known to exist in three alternative forms) is assessedwithin an individual's genome, so that the individual's genotype for thepolymorphism as that site can be completely known. For example,appropriate polymorphisms that can be assessed in the genes listed aboveinclude the following

[0049] i) a polymorphism manifested as occurrence of a codon encodingalanine at amino acid residue 9 (i.e., in the signal sequence) of MnSOD;

[0050] ii) a polymorphism manifested as occurrence of a codon encodingvaline at amino acid residue 9 (i.e., in the signal sequence) of MnSOD;

[0051] iii) a polymorphism manifested as occurrence of a codon encodingisoleucine at amino acid residue 58 of MnSOD;

[0052] iv) a polymorphism manifested as occurrence of a codon encodingthymine at amino acid residue 58 of MnSOD;

[0053] v) a polymorphism manifested as occurrence of a codon encodingvaline at amino acid residue 7 of CZSOD;

[0054] vi) a polymorphism manifested as occurrence of a codon encodingglutamic acid at amino acid residue 7 of CZSOD;

[0055] vii) a polymorphism manifested as occurrence of a codon encodingcysteine at amino acid residue 6 of CZSOD;

[0056] viii) a polymorphism manifested as occurrence of a codon encodingphenylalanine at amino acid residue 6 of CZSOD;

[0057] ix) a polymorphism manifested as occurrence of a cytosine residueat nucleotide residue −262 (i.e., in the promoter region) of thecatalase gene;

[0058] x) a polymorphism manifested as occurrence of a thymine residueat nucleotide residue −262 (i.e., in the promoter region) of thecatalase gene;

[0059] xi) a polymorphism manifested as occurrence of a codon encodingproline at amino acid residue 198 of glutathione peroxidase (hGPX1);

[0060] xii) a polymorphism manifested as occurrence of a codon encodingleucine at amino acid residue 198 of glutathione peroxidase (hGPX1);

[0061] xiii) a polymorphism manifested as occurrence of a codon encodingvaline at amino acid residue 105 of glutathione S-transferase P1(GSTP1);

[0062] xiv) a polymorphism manifested as occurrence of a codon encodingisoleucine at amino acid residue 105 of glutathione S-transferase P1(GSTP1);

[0063] xv) a polymorphism manifested as occurrence of a cytosine residueat nucleotide residue 242 (i.e., in the coding region) of the geneencoding NAD(P)H:quinone oxidoreductase;

[0064] xvi) a polymorphism manifested as occurrence of a thymine residueat nucleotide residue 242 (i.e., in the coding region) of the geneencoding NAD(P)H:quinone oxidoreductase;

[0065] xvii) a polymorphism manifested as occurrence of a thymineresidue at nucleotide residue 113 in exon 3 of the gene which encodesepoxide hydrolase (i.e., resulting in a tyrosine residue in epoxidehydrolase);

[0066] xviii) a polymorphism manifested as occurrence of a cytosineresidue at nucleotide residue 113 in exon 3 of the gene which encodesepoxide hydrolase (i.e., resulting in a histidine residue in epoxidehydrolase);

[0067] xix) a polymorphism manifested as occurrence of an adenineresidue at nucleotide residue −238 (i.e., in the promoter region) of thegene which encodes TNF-alpha (i.e., the TNF-alpha promoter variantdesignated TNF2);

[0068] xx) a polymorphism manifested as occurrence of an adenine residueat nucleotide residue −308 (i.e., in the promoter region) of the genewhich encodes TNF-alpha (i.e., the TNF-alpha promoter variant designatedTNF3);

[0069] xxi) a polymorphism manifested as occurrence of a cytosineresidue at nucleotide residue 242 (i.e., in the coding region) of thephox gene encoding the NADH/NADPH oxidase p22 subunit;

[0070] xxii) a polymorphism manifested as occurrence of a thymineresidue at nucleotide residue 242 (i.e., in the coding region) of thephox gene encoding the NADH/NADPH oxidase p22 subunit;

[0071] xxiii) a polymorphism manifested as a 27 nucleotide residuerepeat in intron 4 (i.e., between nucleotide residues 5130 and 5511) ofthe gene encoding nitric oxide synthase;

[0072] xxiv) a polymorphism manifested as absence of a 27 nucleotideresidue repeat in intron 4 (i.e., between nucleotide residues 5130 and5511) of the gene encoding nitric oxide synthase;

[0073] xxv) a polymorphism manifested as occurrence of an adenineresidue at nucleotide residue −290 (i.e., in the 5′-flanking region) ofthe gene encoding cytochrome P450 (i.e., the polymorphism designated theCYP3A4 cytochrome P450 variant);

[0074] xxvi) a polymorphism manifested as occurrence of a guanineresidue at nucleotide residue −290 (i.e., in the 5′-flanking region) ofthe gene encoding cytochrome P450 (i.e., the polymorphism designated theCYP3A4 cytochrome P450 variant);

[0075] xxvii) a polymorphism manifested as occurrence of a singleguanine residue at nucleotide residue −1607 of the human gene encodingMMP-1; and

[0076] xxviii) a polymorphism manifested as occurrence of a twoconsecutive guanine residues at a site including nucleotide residue−1607 of the human gene encoding MMP-1.

[0077] Occurrence of an individual disorder-associated polymorphism canbe assessed by first contacting a nucleic acid derived from the human'sgenome with a first oligonucleotide that anneals with higher stringencywith the disorder-associated polymorphism than with a correspondingnon-disorder-associated polymorphism and then assessing annealing of thefirst oligonucleotide and the nucleic acid. Annealing of the firstoligonucleotide and the nucleic acid is an indication that the human'sgenome comprises the disorder-associated polymorphism.

[0078] Occurrence of an individual disorder-associated polymorphism canbe further assessed by contacting the nucleic acid with a secondoligonucleotide that anneals with higher stringency with anon-disorder-associated polymorphism than with the correspondingdisorder-associated polymorphism and assessing annealing of the secondoligonucleotide and the nucleic acid. Annealing of the secondoligonucleotide and the nucleic acid is an indication that the human'sgenome comprises the non-disorder-associated polymorphism. By assessingoccurrence of both disorder-associated and non-disorder associatedpolymorphisms in an individual's genome, one can assess whether theindividual is likely homologous for the non-disorder-associatedpolymorphism, homologous for the disorder-associated polymorphism, orheterozygous for the disorder-associated polymorphism andnon-disorder-associated polymorphisms. This information can informselection of an appropriate agent or intervention and an appropriatedose, duration, or intensity of the agent or intervention for improvingskin health or alleviating, inhibiting, or preventing a skin disorder.

[0079] A skin health score can be calculated by summing, for each of theselected genes in which a disorder-associated polymorphism occurs in thehuman's genome, the product of a constant and a correlation factor. Thecorrelation factor can be one or it can, for example, represent thefraction of humans heterozygous or homozygous for thedisorder-associated polymorphism who exhibit the corresponding disorder.The skin health score represents the relative susceptibility of thehuman to a skin disorder.

[0080] The same methods can be used to assess the likelihood that ahuman will develop a skin disorder. Occurrence of any of thedisorder-associated polymorphisms is an indication that the human ismore susceptible to the skin disorder than a human whose genome does notcomprise the polymorphism, and occurrence of a plurality of thedisorder-associated polymorphisms is an indication that the human iseven more susceptible to the skin disorder than a human whose genomedoes not comprise the polymorphisms.

[0081] These methods can also be used to select a dose of a skinprotective composition or other prophylactic or therapeutic compositionfor administration to a human. Occurrence of any of thedisorder-associated polymorphisms is an indication that a greater doseof the composition should be administered to the human. The dose of thecomposition can be selected based on occurrence of the polymorphisms.

[0082] The invention further relates to a kit for assessing relativesusceptibility of a human to a skin disorder. The kit comprises reagentsfor assessing occurrence in the human's genome of disorder-associatedpolymorphisms in at least two (three, four, five, six, eight, ten, orfifteen or more) of the genes disclosed herein.

[0083] In another aspect, the invention relates to a method of assessingthe advisability that a human should employ a nutritional productcomprising a skin protective agent or other prophylactic or therapeuticcomposition. The method comprises assessing occurrence in the human'sgenome of disorder-associated polymorphisms in at least two (three,four, five, six, eight, ten, fifteen, or more) of the genes disclosedherein. Occurrence of any of the disorder-associated polymorphisms is anindication that it is more advisable for the human to employ thenutritional product than a human whose genome does not comprise thepolymorphism, and occurrence of a plurality of the disorder-associatedpolymorphisms is an indication that it is even more advisable that thehuman should employ the nutritional product than a human whose genomedoes not comprise the polymorphisms.

[0084] In still another aspect, the invention relates to a method ofselecting a dose of a skin protective agent for administration to ahuman in a nutritional product. The method comprises assessingoccurrence in the human's genome of disorder-associated polymorphisms inat least two of the genes disclosed herein. Occurrence of any of thepolymorphisms is an indication that a greater dose of the agent shouldbe administered to the human in the nutritional product. The dose of theagent for the nutritional product can be selected based on occurrence ofthe polymorphisms.

BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWINGS

[0085] The foregoing summary, as well as the following detaileddescription of preferred embodiments of the invention, will be betterunderstood when read in conjunction with the appended drawings. Theinvention is not limited to the precise arrangements andinstrumentalities shown.

[0086]FIGS. 1A and 1B are images which depict examples of results thatcan be obtained by analyzing occurrence of polymorphisms in severalgenes. The results shown in FIG. 1A are derived from a hypotheticalfirst human, and those shown in FIG. 1B are derived from a hypotheticalsecond human. Circles represent different polymorphisms of the geneindicated to the left of the row of circles. Filled circles indicate thepresence of the polymorphism. Non-filled circles indicate the absence ofthe polymorphism. Numbers below each circle represent a correlationfactor for the polymorphism and a disease or disorder (i.e., notnecessarily a skin disease or disorder).

DETAILED DESCRIPTION OF THE INVENTION

[0087] The invention relates to kits and methods for assessing skinhealth in a human by assessing occurrence in the human's genome ofgenetic polymorphisms that are associated with disorders (i.e., any typeof disorder, whether a disorder of the skin or not). To bettercharacterize the human's genetic content, occurrence of polymorphicforms (of the same genes) that are not associated with a disorder canalso be assessed, so that one can determine whether the human is 1)homozygous for the disorder-associated polymorphism at a genomic site,2) heterozygous for disorder-associated and non-disorder-associatedpolymorphisms at that site, or 3) homozygous for non-disorder-associatedpolymorphisms at that site. Assessments of genomic polymorphism contentin two or more (and preferably in three, four, five, six, eight, ten,fifteen, or more) of the genes identified herein as being significant toskin health can be combined to indicate the skin health of the human.This assessment of skin health can be used to predict the likelihoodthat the human will develop, is developing, is predisposed to develop,or has already developed a skin disorder.

[0088] Crudely simplified, the methods involve determining whethermultiple polymorphisms that have been associated (by the inventors or byothers) with a human disorder (i.e., a disease or pathological state,whether of the skin or not) occur in the genome of the human beingtested. In some embodiments, the number of polymorphisms that occur inthe human's genome are summed to yield a value; the higher the value is,the greater the susceptibility of the human to skin disorders isassessed to be (i.e., the poorer the human's skin health is assessed tobe). In other embodiments, a weighting factor is assigned to eachpolymorphism tested, and the weighting factors of polymorphisms thatoccur in the human's genome are summed to yield a value that representsrelative skin health (e.g., as assessed by susceptibility to skindisorders). The weighting factor can represent the product of a constantassigned to the gene in which the corresponding polymorphism occurs anda correlation factor that describes how informative occurrence of thepolymorphism is for occurrence of the disorder with which it isassociated. The weighting factor can also be influenced by whether thehuman is homozygous or heterozygous for the disorder-associatedpolymorphism. The invention includes a variety of alternative methodsand kits for performing the methods, as described in greater detailherein.

[0089] Definitions

[0090] As used in this disclosure, the following terms have the meaningsassociated with them in this section.

[0091] A “polymorphism” in a gene is one of the alternative forms of aportion of the gene that are known to occur in the human population. Forexample, many genes are known to exhibit single nucleotide polymorphicforms, whereby the identity of a single nucleotide residue of the genediffers among the forms. Each of the polymorphic forms represents asingle polymorphism, as the term is used herein. Other known polymorphicforms include alternative forms in which multiple consecutive orclosely-spaced, non-consecutive nucleotide residues vary in sequence,forms which differ by the presence or absence of a single nucleotideresidue or a small number of nucleotide residues, and forms whichexhibit different mRNA splicing patterns.

[0092] A “single nucleotide polymorphism” (“SNP”) is one of thealternative forms of a portion of a gene that vary only in the identityof a single nucleotide residue in that portion.

[0093] A “disorder-associated” polymorphism is an alternative form of aportion of a gene, wherein occurrence of the alternative form in thegenome of a human has been correlated with exhibition by the human of adisease or a pathological state, whether the disease or pathologicalstate affects the skin, another tissue, or multiple tissues.

[0094] A “non-disorder-associated” polymorphism is an alternative formof a portion of a gene for which no significant positive correlation hasbeen made between occurrence of the alternative form in the genome andoccurrence of a disease or a pathological state. Non-disorder-associatedpolymorphisms are sometimes designated “neutral” polymorphisms in theart.

[0095] A disorder-associated polymorphism and a non-disorder-associatedpolymorphism “correspond” with one another if the two polymorphisms aretwo alternative forms of the same portion of the gene. By way ofexample, if the identity of residue 100 of a gene is adenine in adisorder-associated polymorphism of the gene and cytosine in anon-disorder-associated polymorphism of the gene, then the twopolymorphisms correspond with one another. It is understood that theremay be three or more corresponding polymorphisms when there are morethan two alternative forms of the same portion of the gene.

[0096] A “characteristic residue” of a polymorphism is a nucleotideresidue, the identity of which is known to vary among the alternativeforms corresponding to the polymorphism.

[0097] A “skin disorder” is a pathological condition characterized bydysfunction, (e.g., inflammation, necrosis, abnormal proliferation,reduced elasticity, defective renewal, irritation, or infection) of someportion of the skin.

[0098] “Skin health” is a measure of the absence of a skin disorder inan individual human (i.e., characterized by normal skin function andappearance) and the likelihood that the individual will continue toexhibit absence of a skin disorder.

[0099] “Toxic oxygen species” include, in approximate order ofdecreasing reactivity, hydroxyl radicals, superoxide radicals, nitricoxide, peroxy nitrite (ONOO-; the product of a reaction between nitricoxide and superoxide radical), and hydrogen peroxide. Ordinary diatomicoxygen is not a toxic oxygen species, as the term is used herein.

[0100] “Oxidative damage” refers to chemical reaction of a normalcellular component (e.g., DNA, a protein, or a lipid) with a toxicoxygen species, whereby at least one normal function of the component isinhibited or eliminated. The terms “oxidative damage” and “oxidativestress” are used interchangeably herein.

[0101] A “molecular beacon oligonucleotide” is a single-strandedoligonucleotides having a fluorescent label (e.g., rhodamine, FAM, TET,VIC, JOE, or HEX) attached to the 5′-end thereof and a fluorescencequencher (e.g., TAMRA or DABCYL) attached to the 3′-end thereof (or viceversa), as described (Kostrikis et al., 1998, Science 279:1228-1229).

[0102] Two molecular beacon oligonucleotides are “spectrally distinct”if they can be differentially detected using spectrophotometric orspectrofluorimetric methods. Examples of characteristics that can beused to differentiate spectrally distinct oligonucleotides includeabsorption or excitation wavelength, emission wavelength, andfluorescent lifetime.

[0103] An “instructional material” is a publication, a recording, adiagram, or any other medium of expression which can be used tocommunicate how to use a kit described herein, numerical values forweighting the significance of various polymorphisms that are detectableusing the kit, or both. The instructional material of the kit of theinvention can, for example, be affixed to a container which contains akit of the invention or be shipped together with a container whichcontains the kit. Alternatively, the instructional material can beshipped separately from the container with the intention that theinstructional material and the kit be used cooperatively by therecipient.

[0104] The “stringency” with which two polynucleotides anneal means therelative likelihood that the polynucleotides will anneal in a solutionas the conditions of the solution become less favorable for annealing.Examples of stringent conditions are known in the art and can be foundin available references (e.g., Current Protocols in Molecular Biology,John Wiley & Sons, N.Y., 1989, 6.3.1-6.3.6). Aqueous and non-aqueousannealing methods are described in that reference and either can beused. In general, a first pair of polynucleotides anneal with higherstringency than a second pair if the first pair is more likely to anneal(or remain annealed) as one or more of the salt concentration,temperature, and detergent concentration are increased.

[0105] With respect to a disorder, a “correlation factor” for adisorder-associated polymorphism is the fractions of humans who areheterozygous or homozygous for the polymorphism who exhibit thedisorder. The correlation factor can, alternatively, be based solely onthose who are heterozygous, solely on those who are homozygous, or onthose who are either heterozygous or homozygous.

[0106] A “non-extendable” nucleotide residue is a nucleotide residuethat is capable of being added to a polynucleotide by a polymerase(i.e., by extension of the polynucleotide in association with acomplement thereof, catalyzed by the polymerase) and that, upon additionto the polynucleotide, renders the polynucleotide incapable of beingfurther extended by the polymerase.

[0107] Description

[0108] The invention relates to kits and methods for assessing the skinhealth of a human by assessing occurrence in the human's genome ofgenetic polymorphisms that are associated with disorders (i.e., skindisorders or other disorders). Unlike other methods that predictsusceptibility to a disorder based on occurrence of a particularpolymorphism in a particular gene, a panel of informative genes andpolymorphisms is disclosed herein. Using two or more of the genes inthis panel, one can assess the susceptibility of a human to a skindisorder, even if the skin disorder has not been specifically associatedwith occurrence of a polymorphism in the panel.

[0109] It has been discovered an individual's skin health can beassessed by determining the polymorphic forms of certain genes that arepresent in the individual's genome. The genes which are assessed aredisclosed herein. Assessment of disorder-associated polymorphisms in twoor more of these genes (preferably three, four, five, six, eight, ten,fifteen, or more of these genes) in a human's genome is predictive ofthe human's skin health. The greater the number of these genes in whichoccurrence of disorder-associated polymorphisms is assessed, the greaterthe precision of the methods for predicting the human's skin health islikely to be. Occurrence in the individual's genome of otherpolymorphisms (e.g., ones known to be associated with occurrence of theskin disorder of interest) can also be assessed concurrently orsequentially.

[0110] Skin disorders for which the kits and methods described hereinare useful include inflammatory disorders (e.g., contact dermatitis,uticaria, atopic dermatitis, psoriasis, lichen planus, cutaneous lupuserythematosus, pemphigus, and scleroderma, sun damage (e.g., reddeningand sun bum), infectious diseases (e.g., bacterial and viralinfections), and skin tumors ( e.g., keratoses, squamous cellcarcinomas, basal cell carcinomas, melanomas, and Kaposi's sarcoma).

[0111] Susceptibility of an individual to a skin disorder can beaffected by oxidative stress that skin cells experience. Several of thegenes having polymorphic forms that are informative for skin healthencode proteins that modulate the body's response to or protection fromoxidative stress. For example, genes which protect against oxidativestress include genes which encode an enzyme that catalyzes conversion ofa toxic oxygen species to a less toxic oxygen species, genes that encodea protein that directly provides protection against oxidative damage,genes which encode a protein that indirectly provides protection againstoxidative damage, genes which encode a component of the human DNA repairsystem, and genes (not necessarily included within the preceding groups)which are associated with inducible production of reactive oxygenspecies in immune cells following microbial infection.

[0112] Polymorphisms have been identified in some, if not all, of thenumerous genes that encode components of the human DNA repair system.Disorder-associated polymorphisms in these genes can be informative forthe skin health of an individual (e.g., for susceptibility of theindividual to a skin disorder). Examples of these genes include thosewhich encode apurinic and apyrimidinic endonucleases, enzymes thatcatalyze excision of nucleotide residues damaged by ultravioletradiation, and enzymes that catalyze site specific-recombination. Manysuch genes are known, and include those listed in Wood et al., 2001,Science 291(5507):1284-1289.

[0113] Skin comprises immune cells and acts as a first line of defenseagainst microbial invasion. Genes that induce production of reactiveoxygen species in immune cells following microbial infection includegenes (e.g., genes which encode components of the humanphagocyte-specific NADPH-oxidase complex) associated with respiratoryburst (sometimes designated oxidative burst) phenomena of macrophagesand polymorphonuclear nucleophilic granulocytes, whereby toxic oxygenspecies are produced in response to invasion of a tissue by a microbe(e.g., a protozoan, or a bacterium such as a Pseudomonas, Salmonella, orSerratia bacterium or a known pathogen such as Bacillus anthracis,Escherichia coli, or Staphylococcus aureus). Also included within thisgroup are genes which are known to be aberrant in patients afflictedwith disorders that inhibit or abolish antimicrobial activities ofmacrophages (e.g., chronic granulomatous disease). Disorder-associatedpolymorphisms in substantially any of these genes can be informative ofthe susceptibility of the individual to a skin disorder, particularly askin infection or inflammatory skin disorder. Identification ofindividuals in whom such polymorphisms occur (e.g., using the methodsdescribed herein) can be used, for example, to assess whether anindividual has an elevated risk for developing a skin disorder andwhether some disorder inhibits intervention should be undertaken.

[0114] It is not critical that the gene in which the occurrence of apolymorphism occurs is recognized as being directly or indirectlyinvolved in a skin disorder. It is sufficient that an association can bemade between either the level of expression of the gene or the sequenceof the gene product and skin health of humans.

[0115] Skin disorders include allergic reactions, such as hives andcontact dermatitis. Genes that encode enzymes that catalyze reactionsresponsible for decreasing electrophilic potential of allergens (ortheir metabolites), a process designated biotransformation of allergens,can affect the skin health of a human. Members of the glutathioneS-transferase (GST) family of enzymes, such as GSTP1, participate in thebiotransformation of allergens. These enzymes also catalyzeinterconversions among reactive forms of oxygen. Occurrence of one ormore polymorphism in one of these GST genes can be used to assess skinhealth of an individual.

[0116] Another protein involved in production of toxic oxygen species bycomponents of the immune system in response to allergen exposure isTNF-alpha. Allen et al. (2000, Immunogenetics 51:201-205) described apolymorphism that occurs at nucleotide residue −308 (i.e., in thepromoter region) of the gene that encodes TNF-alpha. This polymorphismcan be one of those assessed as described herein.

[0117] Among enzymes that catalyze conversion of a toxic oxygen speciesto a less toxic oxygen species, four are of particular relevance, namelymitochondrial MnSOD, cytoplasmic CZSOD, catalase (CAT), and glutathioneperoxidase (GP). Polymorphisms that occur in these genes are known to beassociated with various disorders (see, e.g., Kimura et al., 2000, Am.J. Ophthalmol. 130:769-773). Occurrence of disorder-associatedpolymorphisms in at least one (and preferably two, three, or all) ofthese four genes should be assessed in the methods described herein,given the importance of these genes. Similarly, the kits describedherein preferably include reagents for detecting disorder-associatedpolymorphisms in at least one (and preferably two, three, or all) ofthese four genes. In addition, the significance of occurrence ofdisorder-associated polymorphisms in these genes can be applied byassigning a greater weighting factor to disorder-associatedpolymorphisms of these genes than to disorder-associated polymorphismsin other genes disclosed herein.

[0118] Genes in which disorder-associated polymorphisms occur that areuseful for assessing the skin health of an individual include

[0119] genes which encode an enzyme that catalyzes conversion of a toxicoxygen species to a less toxic oxygen species;

[0120] genes which encode a protein that provides protection againstoxidative stress;

[0121] genes which encode a protein that induces production of a toxicoxygen species;

[0122] genes which encode a protein that indirectly affects oxidativestress;

[0123] genes which encode a protein for which the level of expression ofthe protein is associated with oxidative stress;

[0124] genes which encode a component of the human DNA repair system;and

[0125] genes which encode a protein associated with production of atoxic oxygen species by a macrophage or polymorphonuclear neutrophilicgranulocyte.

[0126] It has been discovered that the following genes are of particularrelevance to skin health:

[0127] a) the gene which encodes mitochondrial MnSOD;

[0128] b) the gene which encodes cytoplasmic CZSOD;

[0129] c) the gene which encodes catalase;

[0130] d) the gene which encodes hGPX1;

[0131] e) the gene which encodes GSTP1

[0132] f) the gene which encodes NAD(P)H:quinone oxidoreductase;

[0133] g) the gene which encodes epoxide hydrolase;

[0134] h) the gene which encodes TNF-alpha;

[0135] i) the phox gene;

[0136] j) the gene which encodes nitric oxide synthase;

[0137] k) the gene which encodes cytochrome P450;

[0138] l) the gene which encodes MMP-1; and

[0139] m) the gene which encodes profilagrin.

[0140] Occurrence in a patient's genome of a disorder-associatedpolymorphism in one of genes a)-l) is an indication that the patient isat a greater risk of developing a skin disorder (or is already afflictedwith the disorder) than a human whose genome does not include thedisorder-associated polymorphism. Occurrence of multipledisorder-associated polymorphisms in these genes in a patient's genomeis an indication that that patient is at greater risk for developing askin disorder (i.e., has poorer skin health) than a human in whosegenome fewer (or none) of the disorder-associated polymorphisms occur.Thus, there is a cumulative effect of disorder-associated polymorphismsin the genes identified herein on the skin health of the human in whichthey occur.

[0141] Occurrence of two copies of the same disorder-associatedpolymorphism in the same human (i.e., homozygosity for thedisorder-associated polymorphism) is an indication that the human is ata greater risk for developing a skin disorder (i.e., has poorer skinhealth) than a human in whom only a single copy of the polymorphismoccurs (i.e., an individual heterozygous for the disorder-associatedpolymorphism). Homozygosity for the disorder-associated polymorphism canbe accounted for by more heavily weighting occurrence of two copies ofthe disorder-associated polymorphism than occurrence of only a singlecopy (e.g., by multiplying the significance associated with occurrenceof the disorder-associated polymorphism by a factor such as two, five,ten or another value).

[0142] Although the invention is not limited to the particulardisorder-associated polymorphisms in the genes identified herein, it isrecognized that disorder-associated polymorphisms that occur inparticular portions of the genes can be more significant indicators ofskin health than disorder-associated polymorphisms that occur inparticular portions of the genes. Thus, disorder-associatedpolymorphisms that occur in the following regions of the indicated genescan be weighted more heavily than disorder-associated polymorphisms thatoccur in other portions of the genes. These polymorphisms include

[0143] A) disorder-associated polymorphisms in the open reading frameencoding mitochondrial MnSOD;

[0144] B) disorder-associated polymorphisms in the open reading frameencoding cytoplasmic CZSOD;

[0145] C) disorder-associated polymorphisms in the promoter region ofthe gene encoding catalase;

[0146] D) disorder-associated polymorphisms in the open reading frameencoding hGPX1;

[0147] E) disorder-associated polymorphisms in the open reading frameencoding GSTP1;

[0148] F) disorder-associated polymorphisms in the open reading frameencoding NAD(P)H:quinone oxidoreductase;

[0149] G) disorder-associated polymorphisms in the open reading frameencoding epoxide hydrolase;

[0150] H) disorder-associated polymorphisms in the promoter region ofthe gene encoding TNF-alpha;

[0151] I) disorder-associated polymorphisms in the open reading frame ofthe phox gene;

[0152] J) disorder-associated polymorphisms in the open reading frameencoding nitric oxide synthase;

[0153] K) disorder-associated polymorphisms in the 5′ flanking region ofthe gene encoding cytochrome P450; and

[0154] L) disorder-associated polymorphisms in the promoter region ofthe gene encoding MMP-1.

[0155] Occurrence of any of a number of particular polymorphisms can beassayed in order to assess an individual's skin health. A non-limitinglist of such polymorphisms include the following:

[0156] I) a polymorphism manifested as a change from an alanine residueto a valine residue at amino acid residue 9 (i.e., in the signalsequence) of mitochondrial MnSOD;

[0157] II) a polymorphism manifested as a change from an isoleucineresidue to a thymine residue at amino acid residue 58 of mitochondrialMnSOD;

[0158] III) a polymorphism manifested as a change from a valine residueto a glutamic acid residue at amino acid residue 7 of cytoplasmic CZSOD;

[0159] IV) a polymorphism manifested as a change from a cysteine residueto a phenylalanine residue at amino acid residue 6 of cytoplasmic CZSOD;

[0160] V) a polymorphism manifested as a change from a cytosine residueto a thymine residue at nucleotide residue −262 (i.e., in the promoterregion) of the catalase gene;

[0161] VI) a polymorphism in the hGPX1 gene manifested as a change froma proline residue to a leucine residue at amino acid residue 198 ofglutathione peroxidase;

[0162] VII) a polymorphism in the GSTP1 gene manifested as a change froma valine residue to an isoleucine residue at amino acid residue 105 ofglutathione S-transferase P1;

[0163] VIII) a polymorphism manifested as a change from a cytosineresidue to a thymine residue at nucleotide residue 242 (i.e., in thecoding region) of the gene encoding NAD(P)H:quinone oxidoreductase;

[0164] IX) a polymorphism manifested as a change from a thymine residueto a cytosine residue at nucleotide residue 113 in exon 3 of the genewhich encodes epoxide hydrolase (i.e., effecting change of from atyrosine residue to a histidine residue in epoxide hydrolase);

[0165] X) a polymorphism manifested as a change to an adenine residue atnucleotide residue −238 (i.e., in the promoter region) of the gene whichencodes TNF-alpha (i.e., the TNF-alpha promoter variant designatedTNF2);

[0166] XI) a polymorphism manifested as a change to an adenine residueat nucleotide residue −308 (i.e., in the promoter region) of the genewhich encodes TNF-alpha (i.e., the TNF-alpha promoter variant designatedTNF3);

[0167] XII) a polymorphism manifested as a change from a cytosineresidue to a thymine residue at nucleotide residue 242 (i.e., in thecoding region) of the phox gene encoding the NADH/NADPH oxidase p22subunit;

[0168] XIII) a polymorphism manifested as a 27 nucleotide residue repeatin intron 4 (i.e., between nucleotide residues 5130 and 5511) of thegene encoding nitric oxide synthase;

[0169] XIV) a polymorphism manifested as a change from an adenineresidue to a guanine residue at nucleotide residue −290 (i.e., in the5′-flanking region) of the gene encoding cytochrome P450 (i.e., thepolymorphism designated the CYP3A4 cytochrome P450 variant); and

[0170] XV) a polymorphism manifested as occurrence of a two consecutiveguanine residues at a site including nucleotide residue −1607 of thehuman gene encoding MMP-1.

[0171] Occurrence of a disorder-associated polymorphism in anindividual's genome can be assessed in any of a variety of ways. In oneembodiment, a simple test (e.g., primer extension, PCR amplification, ormolecular beacon oligonucleotide-binding) is used to determine whetheror not the disorder-associated polymorphism occurs in the individual'sgenome (i.e., without regard to copy number). In another embodiment, atest is used to determine whether the individual's genome includes anon-disorder-associated polymorphism corresponding to a knowndisorder-associated polymorphism in a gene disclosed herein (i.e., as anindication that the individual is at least heterozygous for thenon-disorder-associated polymorphism). In yet another embodiment, a test(i.e., using multiple probes or primers) is used that is able to detectboth disorder-associated polymorphisms and non-disorder-associatedpolymorphisms in two, three, four, or more genes disclosed herein. Usingsuch a test, one can determine both occurrence of a disorder-associatedpolymorphism in an individual's genome and whether the individual ishomozygous or heterozygous for the disorder-associated polymorphism.This test also permits ‘checking’ of results, since it can both accountfor all known polymorphic forms and indicate when a previouslyuncharacterized polymorphism occurs at or near the site of a knownpolymorphism.

[0172] In a kit or method for detecting both disorder-associatedpolymorphisms and non-disorder-associated polymorphisms known for thegenes disclosed herein, one or more (preferably at least two, three,four, five, six, eight, ten, or fifteen or more) of the followingpolymorphisms can be assessed:

[0173] i) a polymorphism manifested as occurrence of a codon encodingalanine at amino acid residue 9 (i.e., in the signal sequence) ofmitochondrial MnSOD;

[0174] ii) a polymorphism manifested as occurrence of a codon encodingvaline at amino acid residue 9 (i.e., in the signal sequence) ofmitochondrial MnSOD;

[0175] iii) a polymorphism manifested as occurrence of a codon encodingisoleucine at amino acid residue 58 of mitochondrial MnSOD;

[0176] iv) a polymorphism manifested as occurrence of a codon encodingthymine at amino acid residue 58 of mitochondrial MnSOD;

[0177] v) a polymorphism manifested as occurrence of a codon encodingvaline at amino acid residue 7 of cytoplasmic CZSOD;

[0178] vi) a polymorphism manifested as occurrence of a codon encodingglutamic acid at amino acid residue 7 of cytoplasmic CZSOD;

[0179] vii) a polymorphism manifested as occurrence of a codon encodingcysteine at amino acid residue 6 of cytoplasmic CZSOD;

[0180] viii) a polymorphism manifested as occurrence of a codon encodingphenylalanine at amino acid residue 6 of cytoplasmic CZSOD;

[0181] ix) a polymorphism manifested as occurrence of a cytosine residueat nucleotide residue −262 (i.e., in the promoter region) of thecatalase gene;

[0182] x) a polymorphism manifested as occurrence of a thymine residueat nucleotide residue −262 (i.e., in the promoter region) of thecatalase gene;

[0183] xi) a polymorphism in the hGPX1 gene manifested as occurrence ofa codon encoding proline at amino acid residue 198 of glutathioneperoxidase;

[0184] xii) a polymorphism in the hGPX1 gene manifested as occurrence ofa codon encoding leucine at amino acid residue 198 of glutathioneperoxidase;

[0185] xiii) a polymorphism in the GSTP1 gene manifested as occurrenceof a codon encoding valine at amino acid residue 105 of glutathioneS-transferase P1;

[0186] xiv) a polymorphism in the GSTP1 gene manifested as occurrence ofa codon encoding isoleucine at amino acid residue 105 of glutathioneS-transferase P1;

[0187] xv) a polymorphism manifested as occurrence of a cytosine residueat nucleotide residue 242 (i.e., in the coding region) of the geneencoding NAD(P)H:quinone oxidoreductase;

[0188] xvi) a polymorphism manifested as occurrence of a thymine residueat nucleotide residue 242 (i.e., in the coding region) of the geneencoding NAD(P)H:quinone oxidoreductase;

[0189] xvii) a polymorphism manifested as occurrence of a thymineresidue at nucleotide residue 113 in exon 3 of the gene which encodesepoxide hydrolase (i.e., resulting in a tyrosine residue in epoxidehydrolase);

[0190] xviii) a polymorphism manifested as occurrence of a cytosineresidue at nucleotide residue 113 in exon 3 of the gene which encodesepoxide hydrolase (i.e., resulting in a histidine residue in epoxidehydrolase);

[0191] xix) a polymorphism manifested as occurrence of an adenineresidue at nucleotide residue −238 (i.e., in the promoter region) of thegene which encodes TNF-alpha (i.e., the TNF-alpha promoter variantdesignated TNF2);

[0192] xx) a polymorphism manifested as occurrence of an adenine residueat nucleotide residue −308 (i.e., in the promoter region) of the genewhich encodes TNF-alpha (i.e., the TNF-alpha promoter variant designatedTNF3);

[0193] xxi) a polymorphism manifested as occurrence of a cytosineresidue at nucleotide residue 242 (i.e., in the coding region) of thephox gene encoding the NADH/NADPH oxidase p22 subunit;

[0194] xxii) a polymorphism manifested as occurrence of a thymineresidue at nucleotide residue 242 (i.e., in the coding region) of thephox gene encoding the NADH/NADPH oxidase p22 subunit;

[0195] xxiii) a polymorphism manifested as a 27 nucleotide residuerepeat in intron 4 (i.e., between nucleotide residues 5130 and 5511) ofthe gene encoding nitric oxide synthase;

[0196] xxiv) a polymorphism manifested as absence of a 27 nucleotideresidue repeat in intron 4 (i.e., between nucleotide residues 5130 and5511) of the gene encoding nitric oxide synthase;

[0197] xxv) a polymorphism manifested as occurrence of an adenineresidue at nucleotide residue −290 (i.e., in the 5′-flanking region) ofthe gene encoding cytochrome P450 (i.e., the polymorphism designated theCYP3A4 cytochrome P450 variant);

[0198] xxvi) a polymorphism manifested as occurrence of a guanineresidue at nucleotide residue −290 (i.e., in the 5′-flanking region) ofthe gene encoding cytochrome P450 (i.e., the polymorphism designated theCYP3A4 cytochrome P450 variant);

[0199] xxvii) a polymorphism manifested as occurrence of a singleguanine residue at nucleotide residue −1607 of the human gene encodingMMP-1; and

[0200] xxviii) a polymorphism manifested as occurrence of a twoconsecutive guanine residues at a site including nucleotide residue−1607 of the human gene encoding MMP-1.

[0201] Another important set of polymorphisms that can be assessed inorder to determine an overall skin health score for a human aredisorder-associated polymorphisms that occur in the human profilagringene. Numerous polymorphic forms of these gene are known, and theassociations of each of these forms with one or more disorders is notyet fully characterized. Of course, whenever a profilagrin polymorphicform is or becomes associated with a disorder, occurrenc of thatdisorder-associated polymorphic form of the profilagrin gene can be usedto assess skin health in a human. Known profilagrin polymorphismsinclude SNPs and filagrin-polymer-length polymorphisms. This latter termrefers to the number of filagrin polypeptides into which the profilagrinpolypeptide is post-translationally cleaved in a human cell. Variousindividual humans are known to harbor profilagrin genes that encode aprofilagrin with at least 9 to 12 filagrin units.

[0202] An important aspect of this invention is that human skin health(e.g., susceptibility to a skin disorder such as psoriasis, eczema, askin cancer, or a bacterial infection) can be associated with occurrencein the human's genome of a disorder-associated polymorphism in one ofthe genes described herein—even if there is no known biochemical orphysiological association between occurrence of the polymorphism andskin health or incidence of a skin disorder. Put another way, thepresent inventors have discovered that genes and polymorphisms disclosedherein are predictive indicators of the state of an individual human'sskin health. By assessing whether or not disorder-associatedpolymorphisms occur in the genes identified herein in an individual (andhow many such polymorphisms occur in those genes), one can assess theindividual's skin health (e.g., as manifested as the likelihood that theindividual has, or will develop a skin disorder).

[0203] If it is determined that an individual has poor skin health(e.g., because multiple disorder-associated polymorphisms occur in theindividual's genome in the genes disclosed herein), then the individualcan be encouraged to make changes to improve their skin health, skinappearance, or to reduce the likelihood of developing skin disorders.Such changes can include use of skin protective compositions (e.g.,nutritional formulas including anti-oxidants, sunscreens, and topical orsystem corticosteroids), use of cosmetic compositions, improvingnutrition, and avoiding sunlight. Determination that an individual hasrelatively poor skin health can also be used as an indication that theindividual should be monitored more closely than others for developmentof skin disorders.

[0204] Early detection of a predisposition to develop a skin disordercan enable an individual (or the individual's physician) to take stepsto delay, inhibit, alleviate (i.e., reduce the severity of), or evenprevent the disorder. The appropriate steps for treating and preventingskin disorders are well known and include modifying diet, exercise, andintake or topical application of nutrients and pharmaceuticals.Palliative, therapeutic, and prophylactic methods are known for manyskin disorders, and these can be undertaken once a patient'ssusceptibility to the disorder is known. Thus, the kits and methodsdescribed herein permit a skin disorder to be treated, inhibited, orprevented. The kits and methods described herein allow theseinterventions to be made at an early stage of the skin disorder (whentreatment is often most effective), or even before the disorder issymptomatically manifested.

[0205] It was not previously appreciated that detection in a human'sgenome of two or more disorder-associated polymorphisms in the genesdisclosed herein is indicative that the human exhibits poorer skinhealth, manifested as greater susceptibility to skin disorders thanindividuals having a genome containing fewer (or none) of thesedisorder-associated polymorphisms. Previous studies are believed to haverecognized only association between a polymorphism in only individualgenes identified herein and a particular disorder. The inventors believethat they are the first to describe methods and kits for assessing ahuman's susceptibility to skin disorders based on occurrence in thehuman of certain polymorphisms that are not recognized as beingassociated with the individual skin disorder.

[0206] It has been discovered that disorder-associated polymorphismsthat occur in the genes identified herein as a)-l) can be used to assessboth an individual's skin health and the likelihood that the individualwill develop (or is currently afflicted with) a skin disorder. In oneembodiment of the kits and method described herein, occurrence ofdisorder-associated polymorphisms (and/or non-disorder-associatedpolymorphisms) is assessed in two or more of the genes identified hereinas a)-l), such as occurrence of a disorder-associated polymorphismsidentified herein as A)-L). By way of example, the kit or method caninvolve assessing occurrence of multiple polymorphisms identified hereinas i)-xxviii).

[0207] Methods of Assessing Skin Health

[0208] The invention includes a method of assessing the skin health(e.g., relative susceptibility to one or more skin disorders) of ahuman. Skin health can be calculated relative to a hypothetical humanwhose genome does not contain a single disorder-associated polymorphismin a gene disclosed herein. Alternatively, susceptibility can becalculated relative to another human who may have one or more differentdisorder-associated polymorphism than the human being assessed. Inpractice, the basis upon which raw susceptibility scores are calculatedis immaterial, so long as the same basis is used for all humans whosescores are to be compared (i.e., so that the scores are relatable to oneanother).

[0209] The relative skin health of a human can be used to assess therisks and benefits of a variety of compositions, conditions, andinterventions. In one embodiment, the skin health of a human can be usedto determine whether the human would benefit by supplementingnutritional intake with a composition that contains one or morevitamins, minerals, or other skin protective agents. Numerous skinprotective agents are known and additional agents are certain to bediscovered over time. The usefulness of the kits and methods disclosedherein does not depend on the identity of the particular agent. Examplesof skin protective agents include vitamins (especially anti-oxidantvitamins), minerals, naturally-occurring amino acids, derivatives ofnaturally-occurring amino acids, plant extracts, and conventional skincare products (e.g., skin softening and moisturizing lotions, Aloeextracts, and the like). Anti-oxidant vitamins are preferablyadministered to skin in a protein-complexed form (e.g., usingpreparations such as the VITAZYME® vitamin preparations sold by ArchPersonal Care Products, L.P. of South Plainfield, N.J.). Similarly, skinprotective minerals such as manganese and selenium are also preferablyadministered to skin in a protein-complexed form (e.g., usingpreparations such as the ACQUA-BIOMIN™ mineral preparations sold by ArchPersonal Care Products, L.P.). Useful skin protective plant extractsinclude gape polyphenols and naturally active botanicals (NABs) such asNAB Pikea robusta (red algae) extract, NAB fennel seed (Foeniculumvulgare) extract, and NAB red clover (Trifollum Pratense) leaf extract.Useful naturally-occurring amino acids and derivatives thereof includeglycine, glutamine, N-acetylcysteine, and trimethylglycine. Furthermore,the skin health, as assessed using a kit or method as described herein,can indicate an appropriate dose of such an agent for an individualpatient.

[0210] The skin protective agent that is administered to an individualsubject can be determined by the overall skin health score, by observingthe genes in which disorder-associated polymorphisms occur, or both.

[0211] For example, if a disorder-associated polymorphism occurs in thesubject's MnSOD gene, then a manganese-containing skin protective agent,a zinc-containing skin protective agent, or a manganese- andzinc-containing skin protective agent (e.g., one of the ACQUA BIOMIN™products) can be applied to the subject's skin to inhibit or alleviateskin disorders.

[0212] If a disorder-associated polymorphism occurs in the subject'sglutathione peroxidase gene, then a skin protective agent comprising oneor more of selenium, grape polyphenols, N-acetylcysteine, glutamine,glycine, or NAB fennel seed can be applied to the subject's skin toinhibit or alleviate skin disorders.

[0213] If a disorder-associated polymorphism occurs in the subject'smicrosomal epoxide hydrolase gene, then a skin protective agentcomprising one or more of N-acetylcysteine, trimethylglycine, ananti-oxidant vitamine (e.g., one of the VITAZYME® products), NAB Pikearobusta, and NAB fennel seed can be applied to the subject's skin toinhibit or alleviate skin disorders.

[0214] If a disorder-associated polymorphism occurs in the subject'stumor necrosis factor-alpha gene, then a skin protective agentcomprising one or both NAB Pikea robusta and NAB red clover leaf can beapplied to the subject's skin to inhibit or alleviate skin disorders.

[0215] Skin health of a human is determined by assessing occurrence inthe human's genome of disorder-associated polymorphisms in a pluralityof genes disclosed herein (e.g., 2, 3, 4, 6, 8, 10, 15, or more genes).Occurrence of a disorder-associated polymorphism in one of these genesis an indication that the human has a greater susceptibility to skindisorders and poorer skin health than a human in whose genome thepolymorphism does not occur. Occurrence of two or more suchpolymorphisms in the human's genome indicates that the human exhibitseven greater susceptibility to skin disorders (and poorer skin health).

[0216] Occurrence of each disorder-associated polymorphism in a genedisclosed herein is not necessarily equally indicative of susceptibilityto skin disorders and poorer skin health. In order to account fordifferences in the significance of various disorder-associatedpolymorphisms, a weighting factor can be assigned to each polymorphismdetected in the methods and kits described herein. As indicated above,some genes have a more significant role in skin health in humans thanothers. Generally, disorder-associated polymorphisms that occur in oneof these genes are more significant than polymorphisms that occur ingenes having less significant roles in skin health. Thus, a greaterweighting factor can be assigned to these polymorphisms than to others.By way of example, the weighting factor assigned to these polymorphismscan be 1 to 10 times greater than the weighting factor assigned todisorder-associated polymorphisms in other genes.

[0217] Another factor which can influence the significance that isassigned to occurrence of a disorder-associated polymorphism in ahuman's genome is the degree to which the polymorphism is correlatedwith the corresponding disorder. Some disorders are highly correlatedwith occurrence of a genetic polymorphism, and other disorders exhibitlower correlation with a polymorphism. When a polymorphism is reportedto be associated with a disorder (i.e., with a disease or pathologicalcondition), a degree of correlation between the polymorphism and thedisorder can be determined or obtained from reports in the literature.One useful way of calculating a factor that describes correlationbetween a polymorphism and a disorder is to calculate an odds ratio thatdescribes the likelihood that an individual in whose genome thedisorder-associate polymorphism occurs will exhibit or develop thedisorder. Because the kits and methods described herein can be used todetect whether the human is homozygous for the disease-associatedpolymorphism, odds ratios calculated for homozygous individuals can alsobe used, if they are available. Odds ratios can be calculated asdescribed in the art.

[0218] For a disorder-associated polymorphism, the odds ratio can becalculated as follows. First, the odds of being afflicted with thedisorder are calculated for a first population in which the polymorphismoccurs by dividing the number of afflicted individuals in the firstpopulation by the total number of individuals in the first population.Second, the odds of being afflicted with the disorder are calculated fora first population in whom the polymorphism does not occur by dividingthe number of afflicted individuals in the second population by thetotal number of individuals in the second population. Third, the oddsratio is calculated by dividing the odds for the first population by theodds for the second population. If the odds ratio is greater than one,then this is an indication that occurrence of the polymorphism isassociated with occurrence of the disorder. Furthermore, the magnitudeof the odds ratio is an indication of the significance of theassociation.

[0219] A skin health score for a human can be determined as follows. Asignificance score can be assigned to each disorder-associatedpolymorphism that is detected in the human's genome using a method orkit described herein. The significance score is a constant (e.g., 1.00),and is multiplied by any significance factor (e.g., 1-10) and by anycorrelation factor that is available. If information is available whichdescribes the correlation between homozygosity for the polymorphism andthe corresponding disorder, then that correlation factor should be usedin place of the correlation factor for mere occurrence of thepolymorphism, at least if the method or kit is used to rule outoccurrence in the subject's genome of correspondingdisorder-non-associated polymorphisms. If significance and correlationfactors are not available, then values of 1.00 should be assigned toeach. The skin health score is determined by summing the significancescore for each disorder-associated polymorphism that is detected usingthe method or kit. This skin health score can be compared with thevalues obtained from other subjects, or it can be compared with thevalue (i.e., zero) which would be expected to occur in a human whosegenome does not include any disorder-associated polymorphism in a genedisclosed herein. A high skin health score corresponds to poor skinhealth. Thus, for two individuals having different skin health scores,the individual having the lower score has better skin health than theindividual having the higher score.

[0220] The method used to assess occurrence of any particulardisorder-associated polymorphism (or disorder-non-associatedpolymorphism) is not critical. Numerous methods of detecting occurrenceof a polymorphism are known in the art, and substantially any of thosemethods can be used in the kits and methods described herein. Naturally,the reagents included in the kit will vary depending on the method to beused to detect the polymorphisms. Examples of some suitable polymorphismdetection methods are provided below.

[0221] In one embodiment, a pair of oligonucleotide primers are used toamplify a portion of the gene that includes a polymorphic region.Detection of one or more of the polymorphisms that occur at thepolymorphic region can be achieved by contacting the amplified portionwith an oligonucleotide having a sequence such that it will anneal understringent conditions with the amplified portion only if one polymorphismoccurs at the portion, but will not anneal with the amplified portion ifanother polymorphism occurs at that portion. Various acceptablestringent conditions are known in the art, and can be modified by theskilled artisan as appropriate to any particular amplifiedportion/oligonucleotide pair. An example of stringent conditions ishybridization in 6×sodium chloride/sodium citrate (SSC) at about 45° C.,followed by one or more washes in 0.2×SSC, 0.1% (w/v) SDS at 50° C.

[0222] In an alternative embodiment, one or more molecular beaconoligonucleotides are used to detect polymorphisms (disorder-associated,non-disorder-associated, or both) in a sample that contains a copy ofthe subject's genome, a fraction of the subject's genome, oramplification products generated from the subject's genome (e.g.,amplified portions of one or more of the genes disclosed herein in whichpolymorphisms are known to occur).

[0223] Molecular beacon probes are single-stranded oligonucleotideshaving a fluorescent label (e.g. rhodamine, FAM, TET, VIC, JOE, or HEX)attached at or near the 5′-end thereof and a fluorescence quencher (e.g.TAMRA or DABCYL) attached at or near the 3′-end thereof (or vice versa),as described (Kostrikis et al., 1998, Science 279:1228-1229). Thesequence of each molecular beacon probe is selected to include twocomplementary hairpin regions, whereby the probe can self-anneal to forma hairpin structure. The 5′-and 3′-ends are brought into closeassociation when the hairpin structure forms. The probe also comprises atargeting portion which is selected to be complementary to a targetsequence (e.g. a single polymorphism of a gene disclosed herein). Thetargeting portion and at least one of the hairpin regions are located inclose proximity to one another, meaning that the targeting portioneither overlaps the hairpin region or flanks it, having no more thanabout 5 nucleotide residues therebetween.

[0224] If the hairpin regions of the molecular beacon probe anneal withone another, then the probe does not fluoresce, because the hairpinstructure forms and the fluorescence quencher attached to one end of theprobe quenches fluorescence of the label attached to the other end ofthe probe. If the targeting portion of the probe anneals with a regionof a nucleic acid having the target sequence, then formation of thehairpin structure is inhibited, the fluorescence quencher is not broughtinto association with the fluorescent label, and the probe fluoresces.Multiple molecular beacon probes can be used in a single reactionmixture, and fluorescence associated with the probes can bedifferentiated if the molecular beacon probes are spectrally distinct.

[0225] Thus, in this embodiment, one or more molecular beacon probes areused, each having targeting portion which is complementary to a targetregion (e.g. 20 to 40 nucleotide residues, more preferably 20 to 30residues) of one polymorphism of a gene disclosed herein. If thepolymorphism to be detected is a single nucleotide polymorphism (SNP),then the target region includes, and preferably is approximatelycentered around, the nucleotide residue at which the polymorphismoccurs. More preferably, two such probes are used, one having atargeting region completely complementary to the target region of onepolymorphism of the gene (e.g., one of two polymorphisms of a particularSNP), and the other having a targeting region completely complementaryto the target region of a corresponding polymorphism of the gene (e.g.,the other polymorphism of the SNP if there are only two polymorphicforms), so that occurrence of disorder-associated andnon-disorder-associated polymorphisms can be simultaneously determined.

[0226] In yet another embodiment of how polymorphisms in gene disclosedherein can be assessed, oligonucleotide primers which are complementaryto a region adjacent a characteristic residue of the polymorphism areextended using a polymerase enzyme, and the identity of the nucleotideresidue that is added to the primer in the position complementary to thecharacteristic residue is determined. The primer can be extended in thepresence of non-extendable nucleotide residues in order to ensure that alimited number of nucleotide residues (or only one) are incorporatedinto the primer. Methods of this type are known in the art (e.g., theSNP-IT® technology of Orchid Biocomputer, Inc.) and are described, forexample in U.S. Pat. Nos. 6,013,431 and 6,004,744.

[0227] Methods of Assessing Susceptibility to Individual Skin Disorders

[0228] An patient's skin health score is predictive of the patient'ssusceptibility to individual skin disorders (a higher score indicating agreater susceptibility to such disorders). The rate or likelihood ofdevelopment and progression of skin disorders can be estimated byassessing the skin health (i.e., determining a skin health score) of apatient. The rate or likelihood of development and progression of theparticular skin disorders disclosed herein can be estimated by assessingoccurrence of the disorder-associated polymorphisms disclosed herein.

[0229] The individual skin disorders for which susceptibility can beassessed using these methods are not limited to those disclosed herein.The methods can be used to assess susceptibility to substantially anyskin disorder. However, it is likely that congenital skin defects whichlead to development of aberrant skin in utero or during the first fewyears of life are unlikely to be associated with the disorder-associatedpolymorphisms described herein.

[0230] Kits for Assessing Skin Health

[0231] The invention includes a kit for assessing the skin health of ahuman and/or the susceptibility of the human to a skin disorder. The kitcontains reagents for performing one or more of the methods describedherein. The reagents used in certain embodiments of the methodsdescribed herein are indicated above. Reagents useful for performingthose methods using a variety of alternative sample preparation andpolymorphism detection methods or chemistries are apparent to theskilled artisan.

[0232] Kits for detecting polymorphisms in individual genes are known inthe art, and the kit of the invention can have similar components.However, a critical feature of the kit is that it includes reagents thatpermit its user to detect disorder-associated polymorphisms in at leasttwo (or at least three, four, six, eight, ten, or fifteen or more) genesdisclosed herein.

[0233] In one embodiment, the kit includes a plurality ofoligonucleotides which anneal under stringent conditions with adisorder-associated polymorphism of one of the genes (e.g., one of thegenes identified herein as being of particular relevance for skinhealth), but not with a non-disorder associated-polymorphism. Each ofthe oligonucleotides can be attached to a surface in order to facilitatehandling of the oligonucleotide. The oligonucleotides can be linked witha plurality of surfaces (e.g., oligonucleotides for a particularpolymorphism being attached to a particle discrete from a particle towhich oligonucleotides for another polymorphism are attached), or theycan be attached to discrete regions of a single surface (e.g., a glassor silicon surface having oligonucleotides attached at defined locationsthereon, as in the GENECHIP™ device of Affymetrix, Inc.). Annealingbetween individual oligonucleotides and the polymorphism correspondingthereto can be detected using standard methods. The kit can alsocomprise oligonucleotides that are useful as molecular beacon probes oras extendable primers.

[0234] In one embodiment, the kit further comprises a DNA collection kitor apparatus, such as that described in co-pending U.S. Pat. No.6,291,171. Advantageously, DNA collected using the kit or apparatus canbe stored or archived, and subjected to additional testing as previouslyunknown disorder-associated polymorphisms are discovered in the genesdisclosed herein, or as the significance of previously unappreciatedpolymorphisms is realized.

[0235] The invention also relates to a method of assessing theadvisability that a human should consume or apply a nutritional productcomprising a skin protective agent such as those described above. Themethod is performed as described herein for assessing the skin health ofa human. If poorer skin health is detected in the human (i.e., relativeto a human not having a disorder-associated polymorphism in a geneidentified herein), then it is more advisable the human should consumeor apply a nutritional product comprising the skin protective agent. Agreater skin health score (i.e., corresponding to poorer skin health) ina human correlates with an increased advisability that the human shoulduse such a nutritional product, and also indicates that a greater doseof the skin agent(s) should be included in the nutritional product.

[0236] It will be appreciated by those skilled in the art that changescan made to the embodiments described above without departing from thebroad inventive concept thereof.

[0237] The disclosure of every patent, patent application, andpublication cited herein is hereby incorporated herein by reference inits entirety.

[0238] This invention is not limited to the particular embodimentsdisclosed, and includes modifications within the spirit and scope of thepresent invention as defined by the appended claims.

What is claimed is:
 1. A method of assessing skin health of a human, themethod comprising assessing occurrence in the human's genome ofdisorder-associated polymorphisms in at least two genes selected fromthe group consisting of a) the gene which encodes mitochondrialmanganese superoxide dismutase (MnSOD); b) the gene which encodescytoplasmic copper/zinc superoxide dismutase (CZSOD); c) the gene whichencodes catalase; d) the gene which encodes human glutathione peroxidase(hGPX1); e) the gene which encodes glutathione S transferase P1 (GSTP1);f) the gene which encodes NAD(P)H:quinone oxidoreductase; g) the genewhich encodes epoxide hydrolase; h) the gene which encodes tumornecrosis factor alpha (TNF-alpha); i) the gene which encodes NADH/NADPHoxidase p22 subunit (the phox gene); j) the gene which encodes nitricoxide synthase; k) the gene which encodes cytochrome P450; l) the genewhich encodes matrix metalloproteinase 1 (MMP-1); and m) the gene whichencodes profilagrin, whereby occurrence of any of thedisorder-associated polymorphisms is an indication that the human haspoorer skin health than a human whose genome does not comprise any ofthe disorder-associated polymorphisms, and whereby occurrence of aplurality of the disorder-associated polymorphisms is an indication thatthe human has even poorer skin health than a human whose genome does notcomprise the disorder-associated polymorphisms.
 2. The method of claim1, wherein occurrence of disorder-associated polymorphisms is assessedin at least three of the genes.
 3. The method of claim 1, whereinoccurrence of disorder-associated polymorphisms is assessed in at leastfour of the genes.
 4. The method of claim 1, wherein occurrence ofdisorder-associated polymorphisms is assessed in at least five of thegenes.
 5. The method of claim 1, comprising assessing occurrence of atleast two disorder-associated polymorphisms selected from the groupconsisting of A) a polymorphism in the open reading frame encodingmitochondrial MnSOD; B) a polymorphism in the open reading frameencoding cytoplasmic CZSOD; C) a polymorphism in the promoter region ofthe gene encoding catalase; D) a polymorphism in the open reading frameencoding hGPX1; E) a polymorphism in the open reading frame encodingGSTP1; F) a polymorphism in the open reading frame encodingNAD(P)H:quinone oxidoreductase; G) a polymorphism in the open readingframe encoding epoxide hydrolase; H) a polymorphism in the promoterregion of the gene encoding TNF-alpha; I) a polymorphism in the openreading frame of the phox gene; J) a polymorphism in the open readingframe encoding nitric oxide synthase; K) a polymorphism in the 5′flanking region of the gene encoding cytochrome P450; and L) apolymorphism in the promoter region of the gene encoding MMP-1.
 9. Themethod of claim 1, comprising assessing occurrence of a firstpolymorphism selected from the group consisting of i) a polymorphismmanifested as occurrence of a codon encoding alanine at amino acidresidue 9 of mitochondrial MnSOD; ii) a polymorphism manifested asoccurrence of a codon encoding valine at amino acid residue 9 ofmitochondrial MnSOD; iii) a polymorphism manifested as occurrence of acodon encoding isoleucine at amino acid residue 58 of mitochondrialMnSOD; iv) a polymorphism manifested as occurrence of a codon encodingthymine at amino acid residue 58 of mitochondrial MnSOD; v) apolymorphism manifested as occurrence of a codon encoding valine atamino acid residue 7 of cytoplasmic CZSOD; vi) a polymorphism manifestedas occurrence of a codon encoding glutamic acid at amino acid residue 7of cytoplasmic CZSOD; vii) a polymorphism manifested as occurrence of acodon encoding cysteine at amino acid residue 6 of cytoplasmic CZSOD;viii) a polymorphism manifested as occurrence of a codon encodingphenylalanine at amino acid residue 6 of cytoplasmic CZSOD; ix) apolymorphism manifested as occurrence of a cytosine residue atnucleotide residue −262 of the catalase gene; x) a polymorphismmanifested as occurrence of a thymine residue at nucleotide residue −262of the catalase gene; xi) a polymorphism manifested as occurrence of acodon encoding proline at amino acid residue 198 of hGPX1; xii) apolymorphism manifested as occurrence of a codon encoding leucine atamino acid residue 198 of hGPX1; xiii) a polymorphism manifested asoccurrence of a codon encoding valine at amino acid residue 105 ofGSTP1; xiv) a polymorphism manifested as occurrence of a codon encodingisoleucine at amino acid residue 105 of GSTP1; xv) a polymorphismmanifested as occurrence of a cytosine residue at nucleotide residue 242of the gene encoding NAD(P)H:quinone oxidoreductase; xvi) a polymorphismmanifested as occurrence of a thymine residue at nucleotide residue 242of the gene encoding NAD(P)H:quinone oxidoreductase; xvii) apolymorphism manifested as occurrence of a thymine residue at nucleotideresidue 113 in exon 3 of the gene which encodes epoxide hydrolase;xviii) a polymorphism manifested as occurrence of a cytosine residue atnucleotide residue 113 in exon 3 of the gene which encodes epoxidehydrolase; xix) a polymorphism manifested as occurrence of an adenineresidue at nucleotide residue −238 of the gene which encodes TNF-alpha;xx) a polymorphism manifested as occurrence of an adenine residue atnucleotide residue −308 of the gene which encodes TNF-alpha; xxi) apolymorphism manifested as occurrence of a cytosine residue atnucleotide residue 242 of the phox gene; xxii) a polymorphism manifestedas occurrence of a thymine residue at nucleotide residue 242 of the phoxgene; xxiii) a polymorphism manifested as occurrence of a 27 nucleotideresidue repeat between nucleotide residues 5130 and 5511 of the geneencoding nitric oxide synthase; xxiv) a polymorphism manifested asnon-occurrence of a 27 nucleotide residue repeat between nucleotideresidues 5130 and 5511 of the gene encoding nitric oxide synthase; xxv)a polymorphism manifested as occurrence of an adenine residue atnucleotide residue −290 of the gene encoding cytochrome P450 (i.e., thepolymorphism designated the CYP3A4 cytochrome P450 variant); xxvi) apolymorphism manifested as occurrence of a guanine residue at nucleotideresidue −290 of the gene encoding cytochrome P450 (i.e., thepolymorphism designated the CYP3A4 cytochrome P450 variant); xxvii) apolymorphism manifested as occurrence of a single guanine residue atnucleotide residue −1607 of the human gene encoding MMP-1; and xxviii) apolymorphism manifested as occurrence of a two consecutive guanineresidues at a site including nucleotide residue −1607 of the human geneencoding MMP-1.
 13. The method of claim 9, comprising assessingoccurrence in the human's genome of disorder-associated polymorphisms inat least four of i) through xxviii).
 14. The method of claim 9,comprising assessing occurrence in the human's genome of at least six ofi) through xxviii).
 15. The method of claim 9, comprising assessingoccurrence in the human's genome of at least ten of i) through xxviii).16. The method of claim 9, comprising assessing occurrence in thehuman's genome of at least fifteen of i) through xxviii).
 17. The methodof claim 9, comprising assessing occurrence in the human's genome ofeach of i) through xxviii).
 18. The method of claim 1, whereinoccurrence of an individual disorder-associated polymorphism is assessedby contacting a nucleic acid derived from the human's genome with afirst oligonucleotide that anneals with higher stringency with thedisorder-associated polymorphism than with a correspondingnon-disorder-associated polymorphism and assessing annealing of thefirst oligonucleotide and the nucleic acid under hybridizationconditions sufficient to differentiate annealing of the firstoligonucleotide with the disorder-associated and non-disorder associatedpolymorphisms, whereby annealing of the first oligonucleotide and thenucleic acid is an indication that the human's genome comprises thedisorder-associated polymorphism.
 19. The method of claim 18, whereinthe first oligonucleotide is attached to a support.
 20. The method ofclaim 19, wherein the support has a plurality of different firstoligonucleotides attached thereto, wherein each oligonucleotide annealswith higher stringency with the disorder-associated polymorphism thanwith a corresponding non-disorder-associated polymorphism of a geneselected from the group.
 21. The method of claim 20, wherein the supporthas attached thereto at least five first oligonucleotides that annealwith higher stringency with the disorder-associated polymorphisms thanwith the corresponding non-disorder-associated polymorphisms.
 22. Themethod of claim 20, wherein the support has attached thereto at leastten first oligonucleotides that anneal with higher stringency with thedisorder-associated polymorphisms than with the correspondingnon-disorder-associated polymorphisms.
 23. The method of claim 20,wherein the support has attached thereto at least fifteen firstoligonucleotides that anneal with higher stringency with thedisorder-associated polymorphisms than with the correspondingnon-disorder-associated polymorphisms.
 24. The method of claim 18,wherein the first oligonucleotide is a molecular beacon oligonucleotide.25. The method of claim 18, wherein occurrence of an individualdisorder-associated polymorphism is further assessed by contacting thenucleic acid with a second oligonucleotide that anneals with higherstringency with a non-disorder-associated polymorphism than with thecorresponding non-disorder-associated polymorphism and assessingannealing of the second oligonucleotide and the nucleic acid underhybridization conditions sufficient to differentiate annealing of thefirst oligonucleotide with the disorder-associated and non-disorderassociated polymorphisms, whereby annealing of the secondoligonucleotide and the nucleic acid is an indication that the human'sgenome does not comprise the disorder-associated polymorphism.
 26. Themethod of claim 25, wherein the second oligonucleotide is attached to asupport.
 27. The method of claim 26, wherein the first and secondoligonucleotides are attached to the same support.
 28. The method ofclaim 25, wherein the second oligonucleotide is a molecular beaconoligonucleotide.
 29. The method of claim 28, wherein the first andsecond oligonucleotides are spectrally distinct molecular beaconoligonucleotides.
 30. The method of claim 1, further comprisingcalculating a skin health score by summing, for each of the selectedgenes in which a disorder-associated polymorphism occurs in the human'sgenome, the product of a constant and a correlation factor, wherein thecorrelation factor represents the fraction of humans heterozygous orhomozygous for the disorder-associated polymorphism who exhibit thecorresponding disorder, whereby the skin health score represents therelative susceptibility of the human to a skin disorder.
 31. The methodof claim 30, wherein the same constant is used for each selected gene.32. The method of claim 1, wherein each of the disorder-associatedpolymorphisms for which occurrence is assessed is a single nucleotidepolymorphism (SNP).
 33. The method of claim 1, wherein occurrence of aSNP is assessed by annealing a nucleic acid derived from the human'sgenome with a primer that is complementary to the region adjacent theSNP on its 3′ side, extending the primer using a polymerase in order toadd a nucleotide residue complementary to the SNP to the primer, anddetecting the identity of the nucleotide residue complementary to theSNP.
 34. The method of claim 33, wherein the nucleotide residue is anon-extendable residue.
 35. A method of assessing the likelihood that ahuman will develop a skin disorder, the method comprising assessingoccurrence in the human's genome of disorder-associated polymorphisms inat least two genes selected from the group consisting of a) the genewhich encodes mitochondrial MnSOD; b) the gene which encodes cytoplasmicCZSOD; c) the gene which encodes catalase; d) the gene which encodeshGPX1; e) the gene which encodes GSTP1; f) the gene which encodesNAD(P)H:quinone oxidoreductase; g) the gene which encodes epoxidehydrolase; h) the gene which encodes TNF-alpha; i) the phox gene; j) thegene which encodes nitric oxide synthase; k) the gene which encodescytochrome P450; l) the gene which encodes MMP-1; and m) the gene whichencodes profilagrin, whereby occurrence of any of thedisorder-associated polymorphisms is an indication that the human ismore susceptible to the skin disorder than a human whose genome does notcomprise the disorder-associated polymorphism, and whereby occurrence ofa plurality of the disorder-associated polymorphisms is an indicationthat the human is even more susceptible to the skin disorder than ahuman whose genome does not comprise the disorder-associatedpolymorphisms.
 36. The method of claim 35, comprising assessingoccurrence of at least two disorder-associated polymorphisms selectedfrom the group consisting of A) a polymorphism in the open reading frameencoding mitochondrial MnSOD; B) a polymorphism in the open readingframe encoding cytoplasmic CZSOD; C) a polymorphism in the promoterregion of the gene encoding catalase; D) a polymorphism in the openreading frame encoding hGPX1; E) a polymorphism in the open readingframe encoding GSTP1; F) a polymorphism in the open reading frameencoding NAD(P)H:quinone oxidoreductase; G) a polymorphism in the openreading frame encoding epoxide hydrolase; H) a polymorphism in thepromoter region of the gene encoding TNF-alpha; I) a polymorphism in theopen reading frame of the phox gene; J) a polymorphism in the openreading frame encoding nitric oxide synthase; K) a polymorphism in the5′ flanking region of the gene encoding cytochrome P450; and L) apolymorphism in the promoter region of the gene encoding MMP-1.
 37. Themethod of claim 35, comprising assessing occurrence of a firstpolymorphism selected from the group consisting of i) a polymorphismmanifested as occurrence of a codon encoding alanine at amino acidresidue 9 of mitochondrial MnSOD; ii) a polymorphism manifested asoccurrence of a codon encoding valine at amino acid residue 9 ofmitochondrial MnSOD; iii) a polymorphism manifested as occurrence of acodon encoding isoleucine at amino acid residue 58 of mitochondrialMnSOD; iv) a polymorphism manifested as occurrence of a codon encodingthymine at amino acid residue 58 of mitochondrial MnSOD; v) apolymorphism manifested as occurrence of a codon encoding valine atamino acid residue 7 of cytoplasmic CZSOD; vi) a polymorphism manifestedas occurrence of a codon encoding glutamic acid at amino acid residue 7of cytoplasmic CZSOD; vii) a polymorphism manifested as occurrence of acodon encoding cysteine at amino acid residue 6 of cytoplasmic CZSOD;viii) a polymorphism manifested as occurrence of a codon encodingphenylalanine at amino acid residue 6 of cytoplasmic CZSOD; ix) apolymorphism manifested as occurrence of a cytosine residue atnucleotide residue −262 of the catalase gene; x) a polymorphismmanifested as occurrence of a thymine residue at nucleotide residue −262of the catalase gene; xi) a polymorphism manifested as occurrence of acodon encoding proline at amino acid residue 198 of hGPX1; xii) apolymorphism manifested as occurrence of a codon encoding leucine atamino acid residue 198 of hGPX 1; xiii) a polymorphism manifested asoccurrence of a codon encoding valine at amino acid residue 105 ofGSTP1; xiv) a polymorphism manifested as occurrence of a codon encodingisoleucine at amino acid residue 105 of GSTP1; xv) a polymorphismmanifested as occurrence of a cytosine residue at nucleotide residue 242of the gene encoding NAD(P)H:quinone oxidoreductase; xvi) a polymorphismmanifested as occurrence of a thymine residue at nucleotide residue 242of the gene encoding NAD(P)H:quinone oxidoreductase; xvii) apolymorphism manifested as occurrence of a thymine residue at nucleotideresidue 113 in exon 3 of the gene which encodes epoxide hydrolase;xviii) a polymorphism manifested as occurrence of a cytosine residue atnucleotide residue 113 in exon 3 of the gene which encodes epoxidehydrolase; xix) a polymorphism manifested as occurrence of an adenineresidue at nucleotide residue −238 of the gene which encodes TNF-alpha;xx) a polymorphism manifested as occurrence of an adenine residue atnucleotide residue −308 of the gene which encodes TNF-alpha; xxi) apolymorphism manifested as occurrence of a cytosine residue atnucleotide residue 242 of the phox gene; xxii) a polymorphism manifestedas occurrence of a thymine residue at nucleotide residue 242 of the phoxgene; xxiii) a polymorphism manifested as occurrence of a 27 basenucleotide residue between nucleotide residues 5130 and 5511 of the geneencoding nitric oxide synthase; xxiv) a polymorphism manifested asnon-occurrence of a 27 nucleotide residue repeat between nucleotideresidues 5130 and 5511 of the gene encoding nitric oxide synthase; xxv)a polymorphism manifested as occurrence of an adenine residue atnucleotide residue −290 of the gene encoding cytochrome P450; xxvi) apolymorphism manifested as occurrence of a guanine residue at nucleotideresidue −290 of the gene encoding cytochrome P450; xxvii) a polymorphismmanifested as occurrence of a single guanine residue at nucleotideresidue −1607 of the human gene encoding MMP-1; and xxviii) apolymorphism manifested as occurrence of a two consecutive guanineresidues at a site including nucleotide residue −1607 of the human geneencoding MMP-1.
 38. A method of selecting a dose of a skin protectivecomposition for administration to a human, the method comprisingassessing occurrence in the human's genome of disorder-associatedpolymorphisms in at least two genes selected from the group consistingof a) the gene which encodes mitochondrial MnSOD; b) the gene whichencodes cytoplasmic CZSOD; c) the gene which encodes catalase; d) thegene which encodes hGPX1; e) the gene which encodes GSTP1; f) the genewhich encodes NAD(P)H:quinone oxidoreductase; g) the gene which encodesepoxide hydrolase; h) the gene which encodes tumor necrosis factor alpha(TNF-alpha); i) the phox gene; j) the gene which encodes nitric oxidesynthase; k) the gene which encodes cytochrome P450; l) the gene whichencodes MMP-1; and m) the gene which encodes profilagrin, wherebyoccurrence of any of the disorder-associated polymorphisms is anindication that a greater dose of the composition should be administeredto the human; and selecting a dose of the composition based onoccurrence of the disorder-associated polymorphisms.
 39. A kit forassessing relative susceptibility of a human to a skin disorder, the kitcomprising reagents for assessing occurrence in the human's genome ofdisorder-associated polymorphisms in at least two genes selected fromthe group consisting of a) the gene which encodes mitochondrial MnSOD;b) the gene which encodes cytoplasmic CZSOD; c) the gene which encodescatalase; d) the gene which encodes hGPX1; e) the gene which encodesGSTP1; f) the gene which encodes NAD(P)H:quinone oxidoreductase; g) thegene which encodes epoxide hydrolase; h) the gene which encodes tumornecrosis factor alpha (TNF-alpha); i) the phox gene; j) the gene whichencodes nitric oxide synthase; k) the gene which encodes cytochromeP450; l) the gene which encodes MMP-1; and m) the gene which encodesprofilagrin.
 40. The kit of claim 39, wherein the reagents comprisefirst oligonucleotides that anneal with higher stringency with thedisorder-associated polymorphisms than with correspondingnon-disorder-associated polymorphisms.
 41. The kit of claim 40, whereineach of the first oligonucleotides is attached to a support.
 42. The kitof claim 41, wherein each of the first oligonucleotides is attached tothe same support.
 43. The kit of claim 41, wherein each of the firstoligonucleotides is attached to a different support.
 44. The kit ofclaim 40, wherein the first oligonucleotides are molecular beaconoligonucleotides.
 45. The kit of claim 40, wherein the kit furthercomprises second oligonucleotides that anneal with higher stringencywith the non-disorder-associated polymorphisms than with correspondingdisorder-associated polymorphisms.
 46. The kit of claim 45, wherein thefirst and second oligonucleotides are spectrally distinct molecularbeacon oligonucleotide pairs.
 47. The kit of claim 39, wherein thereagents comprise primers that are complementary to the region adjacenta characteristic residue of the disorder-associated-polymorphism foramplifying at least the characteristic residue.
 48. The kit of claim 47,further comprising a polymerase capable of extending the primers byadding a nucleotide residue complementary to the characteristic residue.49. The kit of claim 48, further comprising a non-extendable nucleotideresidue.
 50. The kit of claim 39, further comprising an instructionalmaterial which includes a numerical value representing the product of aconstant and a correlation factor, wherein the correlation factorrepresents the fraction of humans heterozygous or homozygous for thedisorder-associated polymorphism who exhibit the corresponding disorder.51. The kit of claim 50, wherein the same constant is used for eachselected gene.
 52. The kit of claim 39, wherein the kit comprisesreagents for assessing occurrence in the human's genome of at least twopolymorphisms selected from the group consisting of I) a polymorphismmanifested as a change from an alanine residue to a valine residue atamino acid residue 9 of mitochondrial MnSOD; II) a polymorphismmanifested as a change from an isoleucine residue to a thymine residueat amino acid residue 58 of mitochondrial MnSOD; III) a polymorphismmanifested as a change from a valine residue to a glutamic acid residueat amino acid residue 7 of cytoplasmic CZSOD; IV) a polymorphismmanifested as a change from a cysteine residue to a phenylalanineresidue at amino acid residue 6 of cytoplasmic CZSOD; V) a polymorphismmanifested as a change from a cytosine residue to a thymine residue atnucleotide residue −262 of the catalase gene; VI) a polymorphism in thehGPX1 gene manifested as a change from a proline residue to a leucineresidue at amino acid residue 198 of glutathione peroxidase; VII) apolymorphism in the GSTP1 gene manifested as a change from a valineresidue to an isoleucine residue at amino acid residue 105 ofglutathione S-transferase P1; VIII) a polymorphism manifested as achange from a cytosine residue to a thymine residue at nucleotideresidue 242 of the gene encoding NAD(P)H:quinone oxidoreductase; IX) apolymorphism manifested as a change from a thymine residue to a cytosineresidue at nucleotide residue 113 in exon 3 of the gene which encodesepoxide hydrolase; X) a polymorphism manifested as a change to anadenine residue at nucleotide residue −238 of the gene which encodesTNF-alpha; XI) a polymorphism manifested as a change to an adenineresidue at nucleotide residue −308 of the gene which encodes TNF-alpha;XII) a polymorphism manifested as a change from a cytosine residue to athymine residue at nucleotide residue 242 of the phox gene; XIII) apolymorphism manifested as a 27 nucleotide residue repeat betweennucleotide residues 5130 and 5511 of the gene encoding nitric oxidesynthase; XIV) a polymorphism manifested as a change from an adenineresidue to a guanine residue at nucleotide residue −290 of the geneencoding cytochrome P450; and XV) a polymorphism manifested asoccurrence of two consecutive guanine residues at a site includingnucleotide residue −1607 of the human gene encoding MMP-1.
 53. A methodof assessing the advisability that a human should employ a nutritionalproduct comprising a skin protective agent, the method comprisingassessing occurrence in the human's genome of disorder-associatedpolymorphisms in at least two genes selected from the group consistingof a) the gene which encodes mitochondrial MnSOD; b) the gene whichencodes cytoplasmic CZSOD; c) the gene which encodes catalase; d) thegene which encodes hGPX1; e) the gene which encodes GSTP1; f) the genewhich encodes NAD(P)H:quinone oxidoreductase; g) the gene which encodesepoxide hydrolase; h) the gene which encodes TNF-alpha; i) the phoxgene; j) the gene which encodes nitric oxide synthase; k) the gene whichencodes cytochrome P450; l) the gene which encodes MMP-1; and m) thegene which encodes profilagrin, whereby occurrence of any of thedisorder-associated polymorphisms is an indication that it is moreadvisable for the human to employ the nutritional product than a humanwhose genome does not comprise the disorder-associated polymorphism, andwhereby occurrence of a plurality of the disorder-associatedpolymorphisms is an indication that it is even more advisable that thehuman should employ the nutritional product than a human whose genomedoes not comprise the disorder-associated polymorphisms.
 54. A method ofselecting a dose of a skin protective agent for administration to ahuman in a nutritional product, the method comprising assessingoccurrence in the human's genome of disorder-associated polymorphisms inat least two genes selected from the group consisting of a) the genewhich encodes mitochondrial MnSOD; b) the gene which encodes cytoplasmicCZSOD; c) the gene which encodes catalase; d) the gene which encodeshGPX 1; e) the gene which encodes GSTP1; f) the gene which encodesNAD(P)H:quinone oxidoreductase; g) the gene which encodes epoxidehydrolase; h) the gene which encodes TNF-alpha; i) the phox gene; j) thegene which encodes nitric oxide synthase; k) the gene which encodescytochrome P450; l) the gene which encodes MMP-1; and m) the gene whichencodes profilagrin, whereby occurrence of any of thedisorder-associated polymorphisms is an indication that a greater doseof the agent should be administered to the human in the nutritionalproduct; and selecting a dose of the agent for the nutritional productbased on occurrence of the disorder-associated polymorphisms.